Alzheimer's disease (AD) is the most common cause of dementia among the elderly. It is estimated that 5.1 million Americans aged 65 or older will develop AD by the year 2010, and the number of people with AD will continue to increase significantly, as the aging population is expected to double over the next 25 years. The Veterans Affairs (VA) health care system serves a large aging population, and because aging is associated with an increased risk of AD, the projected number of aging Veterans at risk for AD is increasing at a dramatic pace; thus, there is a great need for remedies that treat or prevent AD. To date, the only well established susceptibility factor for late-onset Alzheimer's disease (LOAD) is the apolipoprotein E gene (APOE). APOE e4 allele, which is strongly associated with the disease, affects LOAD status and age-at-onset in a dose-dependent manner. However, e4 alone is not necessary or sufficient to cause LOAD. It has therefore been postulated that other genetic, epigenetic, or environmental factors are necessary to compound the APOE e4 effect to develop LOAD. Several lines of evidence, including our own work, suggests the presence of multiple LOAD risk- influencing genetic factors in the APOE locus, and e4 may have a biological function in transcriptional regulation of APOE-locus genes. The proposed studies will explore genetic and biological evidence to demonstrate that APOE e4 has enhancer/silencer activity. This activity can alter the expression profiles of multiple genes in the APOE locus and is functionally relevant for the risk of LOAD. Our short-term goal is to determine the role of e4-related APOE-locus gene regulation in LOAD. Our long-term goal is to precisely define LOAD-causative mechanisms in the APOE region, thereby yielding improved prediction, prevention, and intervention strategies for LOAD. Our hypothesis is that APOE e4 has a strong effect in transcriptional regulation, and that the adverse effects of the e4 in LOAD may be related not only to alterations in apoE protein structure and function but also to changes in the expression of APOE-locus genes. The proposed experiments are specifically aimed at testing this hypothesis. The specific aims of our study will investigate the components of this overall paradigm. Aim 1 will investigate the role of APOE e2/e3/ e4 alleles in the transcriptional regulation of APOE and its adjacent genes. We will use the luciferase reporter gene constructs and assays to study the transcriptional activities in human cell lines. Aim 2 will construct the chromosome-phase-separated haplotypes of the APOE core region, determine the higher- or the lower-risk haplotypes of LOAD, and identify critical LOAD-associated genetic variants. These haplotypes will facilitate haplotype-phenotype correlations of LOAD risk with the transcriptional activities identified in Aim 1. Aim 3 will validate haplotype effects of APOE locus on genes regulation in cellular models and in AD/control postmortem brain.